【摘要】：目的：比較整合素β4(integrin β4, ITGB4)在人肝細(xì)胞癌(Hepatocellular carcinoma, HCC)組織及對應(yīng)的癌旁組織中的表達(dá)差異,比較整合素β4在不同肝癌細(xì)胞系中的表達(dá)差異,分析整合素p4與肝癌臨床病理學(xué)特征之間的聯(lián)系,探討整合素p4在肝癌侵襲轉(zhuǎn)移中的作用。方法：選取華中科技大學(xué)同濟(jì)醫(yī)學(xué)院附屬同濟(jì)醫(yī)院肝臟外科中心手術(shù)切除并經(jīng)病理證實(shí)為肝細(xì)胞癌的手術(shù)標(biāo)本68例,選取具有不同轉(zhuǎn)移能力的肝癌細(xì)胞系7株,利用western-blot,免疫組織化學(xué)(immunohistochemistry, IHC)等方法檢測整合素β4在癌與癌旁組織之間以及不同細(xì)胞系之間的表達(dá)情況,通過統(tǒng)計學(xué)方法分析整合素β4表達(dá)情況對肝癌臨床病理特征的影響。結(jié)果：整合素p4在人肝癌組織中高表達(dá),而在癌旁組織中低表達(dá)或不表達(dá)。癌組織中高表達(dá)的整合素β4與腫瘤的局部侵襲和低分化程度呈正相關(guān)。整合素β4在具有高侵襲轉(zhuǎn)移能力的肝癌細(xì)胞系中(HLF, MHCC97L, MHCC97H, HCCLM3)高表達(dá),而在低侵襲轉(zhuǎn)移能力的肝癌細(xì)胞系中(Hep3B,HepG2,Huh7)低表達(dá)或不表達(dá)。結(jié)論：整合素β4在肝癌組織及具有高轉(zhuǎn)移能力的肝癌細(xì)胞系中高表達(dá),起促進(jìn)肝癌侵襲轉(zhuǎn)移的作用。目的：研究整合素β4(integrin β4, ITGB4)對肝癌細(xì)胞錨著非依賴性生長(anchorage independence)的影響以及調(diào)控該行為的機(jī)制。方法：選擇高表達(dá)整合素β4的肝癌細(xì)胞系HCCLM3和HLF,用RNA干擾(RNAi)技術(shù)敲減整合素β4的表達(dá),利用上述改造后的細(xì)胞系通過軟瓊脂克隆形成實(shí)驗(yàn)(soft agarose colony formation),失巢凋亡分析(anoikis assay)研究整合素β4對肝癌錨著非依賴性生長的影響。通過western blot,免疫共沉淀(Co-Immunoprecipitation, Co-IP),細(xì)胞免疫熒光等技術(shù)研究整合素β4調(diào)控的細(xì)胞信號通路對肝癌細(xì)胞錨著非依賴性生長的影響。結(jié)果：整合素β4增強(qiáng)肝癌細(xì)胞錨著非依賴性生長；整合素β4通過活化AKT/PKB信號通路來促進(jìn)錨著非依賴性生長；整合素β4與表皮生長因子受體(epidermal growth factor receptor, EGFR)以配體非依賴的方式結(jié)合來促進(jìn)錨著非依賴性生長；ITGB4-EGFR復(fù)合物通過FAK (focal adhesion kinase)而不是Src來激活A(yù)KT信號通路。結(jié)論：ITGB4-EGFR復(fù)合物通過活化FAK-AKT信號通路來促進(jìn)肝癌細(xì)胞錨著非依賴性生長。目的：研究整合素β4在體內(nèi)對肝癌生長及肺轉(zhuǎn)移的影響。方法：利用裸鼠皮下成瘤模型研究整合素β4對肝癌在體內(nèi)生長的影響；通過尾靜脈注射肝癌細(xì)胞肺轉(zhuǎn)移模型研究整合素p4對肝癌肺轉(zhuǎn)移的影響。結(jié)果：ITGB4-EGFR活化的FAK-AKT通路可以促進(jìn)肝癌在體內(nèi)生長；ITGB4-EGFR活化的FAK-AKT通路促進(jìn)肝癌肺轉(zhuǎn)移。結(jié)論：ITGB4-EGFR-FAK-AKT信號通路促進(jìn)肝癌體內(nèi)生長及肺轉(zhuǎn)移。
[Abstract]:Objective: to compare the expression of integrin 尾 4 (ITGB4) in human hepatocellular carcinoma (HCC) (Hepatocellular carcinoma, HCC) tissues and corresponding paracancerous tissues, and to compare the expression of integrin 尾 4 in different hepatocellular carcinoma cell lines. To analyze the relationship between integrin p4 and clinicopathological features of hepatocellular carcinoma (HCC) and to explore the role of integrin p4 in the invasion and metastasis of HCC. Methods: 68 cases of hepatocellular carcinoma were selected from the liver surgery center of Tongji Hospital affiliated to Tongji Medical College of Huazhong University of Science and Technology. Seven hepatocellular carcinoma cell lines with different metastatic ability were selected and used western-blot,. Immunohistochemical (immunohistochemistry, IHC) and other methods were used to detect the expression of integrin 尾 4 between cancer and paracancerous tissues as well as between different cell lines. The effect of integrin 尾 4 expression on the clinicopathological characteristics of HCC was analyzed by statistical method. Results: integrin p4 was highly expressed in human hepatocellular carcinoma tissues, but low expression or no expression in paracancerous tissues. The high expression of integrin 尾 4 in cancer tissues was positively correlated with local invasion and low differentiation. Integrin 尾 4 was highly expressed in hepatocellular carcinoma cell line (HLF, MHCC97L, MHCC97H, HCCLM3) with high invasion and metastasis, but low expression or no expression in low invasion and metastasis hepatoma cell line (Hep3B,HepG2,Huh7). Conclusion: the high expression of integrin 尾 4 in HCC and HCC cell lines with high metastatic ability can promote the invasion and metastasis of HCC. Aim: to study the effect of integrin 尾 4 (ITGB4) on the Anchorage independent growth of (anchorage independence) in hepatocellular carcinoma cells and the mechanism of its regulation. Methods: hepatocellular carcinoma cell lines HCCLM3 and HLF, with high expression of integrin 尾 4 were selected to knock down the expression of integrin 尾 4 by RNA interference (RNAi) technique. The modified cell lines were used to form (soft agarose colony formation), by soft Agar cloning. The effect of integrin 尾 4 on Anchor-independent growth of Hepatocellular carcinoma was studied by (anoikis assay) analysis of out-of-nest apoptosis. Western blot, immunoprecipitation (Co-Immunoprecipitation, Co-IP) and cellular immunofluorescence were used to study the effects of integrin 尾 4-regulated cell signaling pathway on anchor-independent growth of HCC cells. Results: integrin 尾 4 enhanced anchor independent growth of HCC cells, and integrin 尾 4 promoted anchor independent growth through activation of AKT/PKB signaling pathway. Integrin 尾 4 binds to epidermal growth factor receptor (epidermal growth factor receptor, EGFR) in a ligand-independent manner to promote anchor-independent growth, and the ITGB4-EGFR complex activates the AKT signaling pathway through FAK (focal adhesion kinase) instead of Src. Conclusion: ITGB4-EGFR complex can promote the growth of HCC cells by activating FAK-AKT signaling pathway. Objective: to study the effect of integrin 尾 4 on the growth and lung metastasis of hepatocellular carcinoma (HCC) in vivo. Methods: the effect of integrin 尾 4 on the growth of hepatocellular carcinoma (HCC) in vivo and the effect of integrin p4 on the lung metastasis of HCC were studied by subcutaneously tumorigenic model of nude mice and the model of lung metastasis of HCC cells injected intravenously. Results: ITGB4-EGFR-activated FAK-AKT pathway could promote the growth of HCC in vivo, and ITGB4-EGFR-activated FAK-AKT pathway could promote lung metastasis of HCC. Conclusion: ITGB4-EGFR-FAK-AKT signaling pathway promotes the growth and lung metastasis of HCC in vivo.
【學(xué)位授予單位】：華中科技大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2016
【分類號】：R735.7

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